Embryonic stem cells (ESCs) have the capacity for unlimited proliferation and differentiation into a wide range of cell types. Our long-term goals are to define the molecular mechanisms underpinning self-renewal and pluripotency of ESCs with the expectation that this information will, (i) define fundamental mechanisms of early embryonic development and, (ii) generate enabling technology that will give utility to ESCs in the area of regenerative medicine. The specific hypothesis is that Myc family transcription factors are important regulators of ESC self-renewal and pluripotency. This is supported by well defined roles for Myc transcription factors in immortalization of tumor cells and roles for the Myc regulatory pathway in early embryonic development. Using both murine and human ESCs, we will determine mechanisms of stem cell self-renewal using biochemical and genetic approaches. The outcomes of this work wil have important implications for general health sciences since ESCs can potentially be used to cure a wide range of degenerative diseases and for repairing chronic injury. Moreover, they are a valuable tool for understanding human embryonic development. The Specific Aims are to: 1. define mechanism(s) by which Myc maintains ESCs in a self-renewing, pluripotent state. This Specific Aim will identify mechanisms of Myc function in self-renewal by characterizing its role in cell cycle control and through the identification of transcriptional targets. This will help elucidate the genetic program underpinning self-renewal and stem cell identity. 2. investigate the relationship between Myc and regulation of self-renewal by the cell cycle machinery. 3. establish mechanisms of self-renewal in human ESCs and to evaluate the role of Myc. Although distinct differences exist between murine and human ESCs, it is likely that common modes of self-renewal regulation will exist. To understand self-renewal of hESCs, we will use information obtained from the mouse ESC model system (Specific Aims 1,2). This information will be crucial for our understanding of human ESC biology and in harnessing their therapeutic potential. The following hESC lines from the NIH Human Embryonic Stem Cell Registry will be used in these studies;BG01, TEO3 and WAO1.